ULTRA-SENSITIVE DETECTION METHODOLOGIES FOR SUBSTANCE P

本研究计划的最终目标是开发一种在高离子生物流体中测量低皮摩尔（pM）浓度P物质（SP）及其代谢物的方法。本提案的第一个目标是开发一种预浓缩和脱盐方法，以允许检测低至亚皮摩尔水平的SP及其代谢物。这将通过评估几种不同类型的固相树脂和膜在生物基质中保留低浓度SP的有效性来完成。本研究的第二个目标是将这种预浓缩/脱盐方法与毛细管电泳（CE）结合使用，并通过柱前或柱后衍生操作激光诱导荧光（LIF）检测。一旦达到这些目标，就有可能通过对大脑进行微透析取样来研究SP在血脑屏障中的转运和代谢。以后的工作将比较开发的CE/LIF方法可实现的检测限与毛细管液相色谱（CLC）与质谱（MS）检测可获得的检测限。未来的研究将致力于构建一个包含预浓缩/脱盐装置、分离毛细管和检测系统的包容性微加工系统。

The ultimate goal of this research proposal is to develop a methodology for the measurement of low picomolar (pM) concentrations of substance P (SP) and its metabolites in highly ionic biological fluids. The first goal of this proposal is to develop a preconcentration and desalting method to permit the detection of low-to-subpicomolar levels of SP and its metabolites. This will be accomplished by evaluating several different types of solid phase resins and membranes for their effectiveness in retaining low concentrations of SP from biological matrixes. The second goal of this research is to use this preconcentration/desalting methodology in conjugation with capillary electrophoresis (CE) with laser-induced fluorescence (LIF) detection operating with either pre or postcolumn derivatization. Once these goals have been met, it will be possible to investigate the transport and metabolism of SP across the Blood Brain Barrier using microdialysis sampling of the brain. Later work will compare the detection limits achievable by the developed CE/LIF methodology to those obtainable by capillary liquid chromatography (CLC) with mass spectrometric (MS) detection. Future studies will work toward the construction of an inclusive microfabricated system that contains the preconcentration/desalting device, separation capillary, and detection system.